Altered indirect hemagglutination method for easy serotyping of Haemophilus parasuis / Método de Hemaglutinação Indireta alterado para facilitar a sorotipificação de Haemophilus parasuis

Glässer's disease is an emergent bacterial disease that affects swine husbandries worldwide causing important economic losses. The aetiological agent, Haemophilus parasuis, is currently divided in fifteen serovars but an increasing number of non-typeable serovars have been reported. Indirect hemagglutination (IHA) is indicated as a serotyping method for H. parasuis. In the present study, we describe an additional step that aims to work around a possible obstacle in the original protocol that may compromise the outcome of this assay. We observed that the choice of anticoagulant for blood collection influences and/or impairs spontaneous adsorption of H. parasuis antigens on sheep red blood cells (SRBCs). However, regardless of the anticoagulant used, chemical treatment of SRBCs with tannic acid induces a stable antigen adsorption (sensitization step). The addition of 1% BSA to SRBCs washing buffer and to antisera dilution augments IHA specificity. Tannic acid treated SRBCs combined with thermo-resistant H. parasuis antigens increases the assay resolution. Thus, our results demonstrate an improvement in the technique of H. parasuis serotyping that will prove valuable to understand Glässer's disease epidemiology and to better characterize serovars involved in outbreaks.(AU)

A Doença de Glässer é uma doença bacteriana emergente que afeta a produção de suínos em todo o mundo e causa importantes perdas econômicas. O agente etiológico, Haemophilus parasuis, é atualmente dividido em quinze sorovares; no entanto, um número crescente de cepas não tipificáveis tem sido relatado. O teste de hemaglutinação indireta (IHA) tem sido utilizado para a sorotipificação de H. parasuis. Neste estudo, descrevemos uma alteração no protocolo original de IHA e que supera uma limitação específica que pode comprometer o uso geral deste ensaio. Descobrimos que o tipo de anticoagulante utilizado para coletar os eritrócitos ovinos (SRBCs) pode comprometer a adsorção espontânea dos antígenos do H. parasuis. Por outro lado, o tratamento químico dos SRBCs com ácido tânico promove uma adsorção antigênica estável (passo de sensibilização) e independente do anticoagulante utilizado. O uso de 1% de BSA durante as lavagens dos SRBCs e na diluição dos antissoros incrementa a especificidade da IHA e, a combinação dos SRBCs tratados quimicamente com antígenos de H. parasuis termo-resistentes aumentam a resolução da IHA. Nossos resultados destacam uma melhoria na principal técnica de sorotipificação de H. parasuis, que auxiliará diretamente no entendimento da epidemiologia da Doença de Glässer e na caracterização dos sorovares envolvidos em surtos da doença.(AU)

Single Center Experience of Five Diffuse Panbronchiolitis Patients Clinically Presenting as Severe Asthma

Diffuse panbronchiolitis (DPB) is a bronchiolitis affecting the whole lung fields which can be treated by macrolide. Especially East Asian patients are more susceptible to diffuse panbronchiolitis. As asthma and DPB both can cause airway obstruction, differential diagnosis is important for the 2 diseases. Here we report 5 patients with DPB clinically presenting as severe asthma in Korea, who were well treated by macrolide. Among the 5 patients, 2 could stop their asthma inhalers and the other 3 could reduce asthma medications after diagnosis and treatment of DPB. In conclusion, considering DPB as differential diagnosis for asthmatics in Asian ethnic groups is important.

Absceso cerebral por Haemophilus influenzae serotipo e en un paciente pediátrico con Síndrome de Apert / Brain abscess caused by Haemophilus influenzae type e in a pediatric patient suffering from Apert syndrome

Se presenta el caso de un absceso cerebral causado por Haemophilus infl uenzae tipo e, en un paciente de 12 años con síndrome de Apert. El síndrome de Apert se caracteriza por el cierre prematuro de las suturas craneales. En 2010, el paciente presentó traumatismo craneano en región frontal, fractura y fístula de líquido cefalorraquídeo. En febrero de 2013 consultó por fi ebre, vómitos y convulsión tónica clónica generalizada, con deterioro progresivo del sensorio. La tomografía axial computarizada mostró una lesión frontal derecha, edema perilesional, leve dilatación ventricular y pansinusitis. Se diagnosticó absceso cerebral con pioventriculitis y se realizó drenaje. Se obtuvo desarrollo de un cocobacilo gram negativo, que fue identifi cado como H. infl uenzae serotipo e. Se realizó tratamiento empírico con meropenem (120 mg/kg/día) y vancomicina (60 mg/kg/día). Luego del resultado del cultivo, se rotó a ceftriaxona (100 mg/kg/día) y metronidazol (500 mg/8 h). El paciente cumplió 8 semanas de tratamiento y se observó evolución favorable

We report a case of a brain abscess caused by Haemophilus infl uenzae type e in a 12 year-old patient suffering from Apert syndrome. Apert syndrome is characterized by the premature closure of cranial sutures. In 2010 the patient suffered head trauma in the frontal area with cranial fracture and a cerebrospinal fl uid fi stula. In February 2013 he was admitted to hospital with fever, vomiting and generalized tonic-clonic seizure with deteriorating mental status/progressive sensory impairment. The computerized axial tomographic scan showed a right frontal lesion, perilesional edema, mild ventricular dilatation and pansinusitis. A brain abscess was diagnosed and drained. The clinical sample was then cultured. A gram negative coccobacillus was isolated and identifi ed as Haemophilus infl uenzae serotype e. Empirical treatment was started with meropenem (120 mg/kg/day) and vancomycin (60 mg/kg/day), which was later switched to ceftriaxone (100 mg/kg/day) and metronidazole (500 mg/8 h) after culture results arrived. The patient was discharged in good clinical condition

Community-acquired pneumonia in the childhood: analysis of the diagnostic methods

Immunological assays such as CIE, LA, and Dot-ELISA were compared in order to diagnose community-acquired pneumonia. Serum, pleural fluid and urine samples were comparatively employed for bacterial antigen detection. Dot-ELISA proved to be an original and practical alternative procedure for detecting bacterial polysaccharide antigens from pleural fluid and/or concentrated urine samples, providing a rapid diagnosis for pediatric patients with community-acquired pneumonia.

Haemophilus aphrophilus brain abscess in the first decade.

This report presents a case of brain abscess due to Haemophilus aphrophilus in a six-year old boy. He was admitted to our hospital suffering from left-sided weakness. The initial radiological diagnosis was an intracranial abscess. Purulent material was obtained by puncturing the subcortical lesion and the sample was cultured on conventional media. H. aphrophilus was isolated in pure culture, identified according to conventional methods and confirmed by Becton Dickinson Laboratory (BBL) crystal system. After surgical drainage and eight weeks of antibiotic therapy, the neurological findings improved. The presented case is an example of H. aphriphilus causing brain abscess in the first decade without cardiac predisposition and with good outcome.

Detection Rates of Bacteria in Chronic Otitis Media with Effusion in Children

This study was performed to investigate polymerase chain reaction-based detection of bacterial DNA in middle ear fluid and assess the correlation between the PCR-positive rate with several factors associated with middle ear effusion. The purpose was to gain a further understanding of bacterial infection as a major cause of otitis media with effusion. Of the 278 specimens of middle ear fluid, 39 (14%) tested positive by ordinary culture. The overall detection rate of bacterial DNA using the PCR method was 36.7% for middle ear effusion, and bacterial DNA detection rates of Hemophilus influenzae, Streptococcus pneumoniae, Moraxella catarrhalis in the middle ear effusion were 29.1%, 4.7% and 10.8%, respectively. The bacterial DNA detection rate was higher in ears with a history of acute otitis media than those without the history. High detection rates were observed in patients younger than 48 months who have had a higher tendency to present with acute otitis media. We concluded that PCR is a more sensitive method for the detection of bacteria in middle ear effusion than ordinary culture, and acute otitis media is a major contributor to the pathogenesis of otitis media with effusion.

Comparison of disc diffusion & E test methods with agar dilution for antimicrobial susceptibility testing of Haemophilus influenzae.

BACKGROUND & OBJECTIVES: Reliable methods of detection of antimicrobial resistance are of paramount importance in the treatment and management of infections caused by Haemophilus influenzae. The objective of the present study was to compare and evaluate the performance of disc diffusion and E test (Epsilometer test) with agar dilution method for antimicrobial susceptibility testing of H. influenzae. METHODS: A total of 46 isolates of H. influenzae from various invasive sites were included as test strains. Antimicrobial susceptibility testing was performed by Kirby-Bauer disc diffusion method for ampicillin, chloramphenicol, trimethoprim-sulphamethoxazole (TMP-SMZ) and cefotaxime. Minimum Inhibitory Concentration (MIC) determination was performed by E test and agar dilution for the same set of antimicrobials. All tests were performed on Haemophilus test medium (HTM). RESULTS: Disc diffusion showed a very major (2%) and minor (4%) interpretative error with TMP-SMZ and minor interpretative errors to ampicillin (13%) and chloramphenicol (24%) when compared to agar dilution method. E test produced only minor interpretative errors to chloramphenicol (7%) and TMP-SMZ (2%) and no interpretative errors with ampicillin and cefotaxime as against agar dilution. E test showed good agreement with agar dilution for each of the antimicrobial tested. INTERPRETATION & CONCLUSION: Disc diffusion test may be used as a preliminary screen for susceptibility testing of H. influenzae. E test is simple, easy to perform and a reliable method for determination of resistance in H. influenzae. However its cost and limited availability in India may limit its use. The reference agar dilution method can be used reliably in routine susceptibility testing of H. influenzae.

Nasopharyngeal carriage of Haemophilus influenzae.

OBJECTIVE: Nasopharyngeal colonization of Haemophilus influenzae (H. influenzae) in young children may be important in developing countries. METHOD: In this study, we screened school going children for carriage of H. influenzae. A total of 44 H. influenzae isolates out of a collection of 162 were characterized for biotypes, capsular serotypes and antibiotic resistance. RESULTS: A significant proportion of H. influenzae (25/44) isolates were serotype b. High antibiotic resistance was observed against commonly administered antibiotics like ampicillin (79%), chloramphenicol (20%), trimethoprim sulfamethoxazole (84%) and erythromycin (95%). Comparison of antibiotic resistance profile of nasopharyngeal isolates was observed to be correlated with those of H. influenzae from disease. CONCLUSION: Multidrug resistant nasopharyngeal H. influenzae in young healthy children may act as reservoir. Monitoring of antibiotic resistance among nasopharyngeal H. influenzae as a surrogate for invasive H. influenzae seems an attractive option.

Rapid detection of Haemophilus influenzae type b in Bangladeshi children with pneumonia and meningitis by PCR and analysis of antimicrobial resistance.

A polymerase chain reaction (PCR) assay with primers from 'bexA' gene was compared with culture for the detection of Haemophilus influenzae type b (Hib) in clinical samples from children with pneumonia and meningitis. Of 200 sera (180 from pneumonia, 20 from non-pneumonia patients) tested by PCR (serum-PCR), Hib was detected in 15 of 16 blood culture-positive and in 6 blood culture-negative pneumonia cases. When compared with the results of blood culture, serum-PCR had sensitivity, specificity, and accuracy index of 93.7%, 96.7%, and 96.5% respectively. Of 120 cerebrospinal fluid (CSF) samples from meningitis patients tested by culture and PCR (CSF-PCR), the latter method could detect Hib in all 15 culture-positive and in 8 of 105 culture-negative cases, showing sensitivity, specificity, and accuracy index of 100%, 92.4%, and 94.4% respectively. The PCR result was available within a day. Antimicrobial susceptibility of Hib was determined by the disc-diffusion method. High rate of resistance to ampicillin (54.8%), chloramphenicol (48.4%), and co-trimoxazole (80.6%) was observed among 31 invasive Hib isolates with resistance to all 3 drugs (multiresistance) in 48.4% of the isolates. All the Hib isolates were susceptible to ceftriaxone. The study has shown that PCR is a rapid, sensitive and specific diagnostic test for Hib from clinical samples, and a combination of culture and PCR is necessary for the detection of Hib infections to the maximum extent for case management to reduce morbidity, mortality, and complications of the invasive Hib infections. A high prevalence of multiresistant Hib strains is a matter of concern.

Verification of the presence of capsule gene sequences in nasopharyngeal isolates of nontypeable Haemophilus influenzae from healthy children at a brazilian day care center

Fifty-eight nasopharyngeal isolates of Haemophilus influenzae were collected from healthy children at a day care center, and nontypeable isolates were examined by Southern blot for the presence of capsule gene sequences. Seven isolates (12per cente) demonstrated homology with capsule-specific sequences. One isolate was characterized as an H. influenzae type b capsule-deficient strain.

Infecciones invasivas por haemophilus influenzae: análisis clínico y factores de riesgo de mortalidad / Haemophilus influenzae invasive infections: clinical characteristics and risk factors of mortality

Durante el período comprendido entre el 1/1/89 y el 31/12/94 se analizaron en forma retrospectiva 220 pacientes con infecciones invasivas por Haemophilus influenzae internados en el Hospital J.P. Garrahan. L a media de edad fue de 11 meses. El 83 por ciento (182p) eran menores de 2 años. La localización meníngea fue la más frecuente. Cuarenta cepas (18.6 por ciento) fueron resistentes a la ampicilina y 2.3 por ciento (5 cepas) al cloranfenicol. El 100 por ciento de las cepas fueron sensibles a cefalosporinas de tercera generación. La mortalidad global fue del 6 por ciento (13 pacientes). Al analizar los factores de riesgo de mortalidad a través del análisis de regresión logística múltiple en 175 pacientes de los que se conocían todas las variables a analizar, la edad menor de un año fue la única variable estadísticamente significativa. Se confirma que el riesgo de morir aumenta a medida que la edad disminuye, independientemente del resto de las variables.

Efusión pleural paraneumónica: experiencia clínica de seis años / Parapneumonic pleural effusion: 6 year clinical experience

Se presenta una revisión de 109 casos de efusión pleural paraneumónica tratados en el Hospital Dr. Exequiel González Cortés durante un período de 6 años (enero 1991 a diciembre 1996). Se clasificaron de acuerdo al aspecto macroscópico y al estudio bacteriológico del líquido pleural, en efusión serosa (21,1 por ciento), fibrinopurulenta o empiema (78,0 por ciento) y organizada (0,9 por ciento). El manejo de los casos de efusión serosa fue sólo con antibióticos en el 69,6 por ciento y en 30,4 por ciento se instaló, además, un tubo pleural. Los pacientes con efusión fibrinopurulenta se manejaron con tubo pleural en el 91,8 por ciento de los casos y un 22,4 por ciento de ellos requirió tratamiento quirúrgico adicional. Se discute el manejo de la efusión pleural, el que continúa siendo controvertido, la utilidad del análisis del líquido pleural para categorizar precozmente la severidad de la enfermedad y la oportunidad de nuevos tratamientos, como cirugía toracoscópica. Se propone un esquema de manejo de los casos, incorporando este nuevo procedimiento terapéutico

Systemic Haemophilus influenzae disease in Thai children.

Fifty patients with systemic Haemophilus influenzae disease were indentified by hospital chart review between 1980-1992. The age distribution varied from 8 days to 14 years; the mean age of the patients was 12.7 months. The peak incidence was between 4 and 6 months of age. There were 27 male patients and 23 female patients for a male:female ratio of 1.17:1. The relative frequencies of 79 clinical entities encountered in 50 patients are as follows: meningitis 55.7%, bacteremia 13.9%, pneumonia 25.3%, cellulitis 2.5% arthritis 1.3%, septic shock 1.3%. There were 23 patients (46%) who had more than one disease entity. Most of the patients were anemic (Hb < 10 gm%) when hospitalization. Sixty-four percent of the patients had early complications. The mortality rate was 8%. Although serotyping was not done from the isolates, at least 33 cerebrospinal fluid samples were positive for H. influenzae type b capsular antigen by counterimmunoelectrophoresis. The percentage of susceptible H. influenzae to penicillin, ampicillin, chloramphenicol, co-trimoxazole were 57.1%, 76.4%, 87.5% and 54.2%, respectively. There was no strain resistant to third generation cephalosporin. Our data indicate that H. influenzae is a serious and life threatening infection. Early diagnosis and proper treatment will reduce the morbidity and mortality rates. For prevention of infection, an appropriate strategy for vaccination is required.

O valor preditivo positivo do esfregaço de Papanicolaou no diagnóstico da vaginite por Gardnerella vaginallis / The positive predictive value of the Pap smear on diagnosis of gardnerella vaginalis vaginitis

Com o objetivo de se avaliar a importância da presença de "células indicadoras" para Gardnerella vaginalis no esfregaço de Papanicolaou no diagnóstico por Gardnerella vaginalis, foi realizado um estudo entre 450 pacientes. Entre essas, 77 (17,6 por cento) eram portadoras de "células indicadoras". Em 35 destas pacientes foi realizada a cultura bacteriológica para Gardnerella vaginalis, confirmando-se o diagnóstico em 31 (88,6 por cento). Nossos resultados mostraram, portanto, uma correlaçäo muito boa entre a presença de "células indicadoras" em esfregaços de Papanicolaou e a cultura de Gardnerella vaginalis

Incidence of Gardnerella vaginalis infection in pregnant & non-pregnant women with non-specific vaginitis.

The occurrence of G. vaginalis in patients with non-specific vaginitis (NSV) was studied. Of the 200 women with vaginitis screened, 84 were diagnosed to have non-specific vaginitis. G. vaginalis was isolated from 33 (39.28%) patients either alone or in combination with other organisms, while one of the healthy controls with no vaginal discharge yielded G. vaginalis on culture. The nature of vaginal discharge could not be correlated with the type of infection. Clue cells and amine test gave inconsistent results in the presence of G. vaginalis infection. A varying susceptibility pattern of G. vaginalis was observed against various antimicrobials, metronidazole (95%) and gentamicin (75%) showing highest sensitivity.

Diagnóstico laboratorial e importância clínica da Gardnerella vaginalis em pacientes com infecçäo vaginal / Laboratory diagnosis and clinical importance of Gardnerella vaginalis in patients with vaginal infections

Foram estudados fluxos cérvico-vaginais de 145 pacientes ambulatoriais com idades entre 16 e 52 anos. Sessenta e cinco pacientes, que apresentavam sintomas clínicos de infecçäo vaginal foram enquadrados no grupo das sintomáticas. Setenta e seis pacientes sem infecçäo genital, aparente, constituiram o grupo controle, onde näo houve isolamento de Gardnerella vaginalis. Das 65 pacientes sintomáticas 19 (29%) apresemtaram G. vaginalis no seu trato genital, 12 (18%) Trichomonas vaginalis e 7 (11%) Candida albicans. Apenas em um dos casos houve associaçäo de G. vaginalis e T. vaginalis. Neste estudo a G. vaginalis foi o mais importante patógeno isolado de mulheres com infecçäo vaginal

Difusión de H. influenzae tipo B (HI B) en contactos intrafamiliares de niños con infecciones sistémicas por este agente / Spread of haemophilus type B in household contacts of patients with invasive infections

Actualmente está bien documentada la difusibilidad de Hi b especialmente en niños y se ha planteado quimioprofilaxis a los menores de 4 años que son contactos de infecciones sistémicas. Para conocer nuestra situación epidemiológica, se estudiaron 31 infecciones sistémicas por Hi b en pacientes ingresados a los hospitales L. Calvo Mackenna y Exequiel González Cortés (24 meningitis, 3 celulitis faciales, 1 neumonía abscesada, 3 pleuroneumonías, una de las cuales presentó además pericarditis) y se investigó la presencia de portadores entre los contactos intrafamiliares. Se estudiaron 191 contactos (X 6,4 por cada caso índice), encontrándose 18% de portadores entre los niños y 5,9% entre los adultos. Se detectó un caso secundario en un contacto de 7 meses de edad. Se logró la erradicación de todos los portadores mediante tratamiento con rifampicina durante 4 días. Se determinó el perfil de sensibilidad a los antimicrobianos de las cepas de Hi aisladas de pacientes y portadores, observándose 12% de cepas resistentes a ampicilina y cloramfenicol entre las cepas de los casos índices. No se observó resistencia en Hi b aislados de portadores